Cryptic species diversity in a widespread bumble bee complex revealed using mitochondrial DNA RFLPs

Abstract

Cryptic species diversity is thought to be common within the class Insecta, posing problems for basic ecological and population genetic studies and conservation management. Within the temperate bumble bee (Bombus spp.) fauna, members of the subgenus Bombus sensu stricto are amongst the most abundant and widespread. However, their species diversity is controversial due to the extreme difficulty or inability morphologically to identify the majority of individuals to species. Our character-based phylogenetic analyses of partial CO1 (700 bp) from 39 individuals spread across their sympatric European ranges provided unequivocal support for five taxa (3-22 diagnostic DNA base pair sites per species). Inclusion of 20 Irish specimens to the dataset revealed ! 2.3% sequence divergence between taxa and 1.3% within taxa. We developed a PCR-RFLP based method for unequivocally distinguishing amongst the four cryptic European taxa of this subgenus, B. cryptarum, B. lucorum, B. magnus and B. terrestris, and used it to analyse 391 females of the former three species collected across Ireland, all of which could be unambiguously assigned to species. Bombus lucorum was the most widely distributed and abundant of the cryptarum-lucorum-magnus species complex, comprising 56% of individuals, though it was significantly less abundant at higher altitudes (>200 m) whilst B. cryptarum was relatively more abundant at higher altitudes. Bombus magnus was rarely encountered at urban sites. Both B. lucorum and B. terrestris are nowadays reared commercially for pollination and transported globally. Our RFLP approach to identify native fauna can underpin ecological studies of these important cryptic species as well as the impact of commercial bumble bees on them.

Revealing the hidden niches of cryptic bumblebees in Great Britain: Implications for conservation

“…B. magnus and B. cryptarum have been regarded as subspecies of B. lucorum and are often referred to collectively as the 'lucorum complex' or simply synonymized to B. lucorum (Benton, 2006;Edwards and Jenner, 2005). Recent studies using CO1 barcode analysis show discrete differences between the three species Murray et al, 2008;Williams et al, 2012), in accordance with studies of labial gland secretions (Bertsch et al, 2005). Diagnostic morphological characters have also been previously reported for queens, but some of these have now been demonstrated to overlap considerably, and vary along a continuum, thus making them unreliable and leading to a high potential for misidentification .…”

“…In July 2010, 13 locations were sampled along a North-South line through the approximate centre of Scotland and England; during June-August 2011, 14 further locations were sampled focussing on the periphery of the UK. The 2011 fieldwork tested the hypothesis that B. magnus is a heathland specialist (Murray et al, 2008;Waters et al, 2010) using a paired sampling design: 11 of the 14 locations comprised a pair of sites representing heathland and non-heathland habitats within 15 km of one another. All locations sampled in 2010 consisted of non-heathland habitat, although some were close to heathland.…”

“…For species identification we followed a PCR-RFLP method based on amplification of the cytochrome oxidase I (COI) gene developed by Murray et al (2008). The pattern of digested fragments for each individual was compared with the characteristic patterns associated with each of the cryptic species and B. terrestris (see Fig.…”

“…The difficulty in identifying these species means that little is known about their ecological attributes; much of what can be found in standard texts will actually be referring to data for multiple species and is therefore of limited value. Consequently, the only reliable information we have on the ecology of these three species comes from Murray et al (2008) and Stanley et al (2013) who used molecular methods to study the lucorum complex in Ireland and Waters et al (2010) who studied them in the Western Isles of Scotland. Niche-partitioning might be expected between these species (Goulson et al, 2008b) and indeed some ecological differences have been suggested.…”

Identification of Bombus species based on wing venation structure

“…Despite considerable intraspecific variation, studies based on morphology (Krüger 1958;Pekkarinen et al 1979;Rasmont 1984), cross-breeding experiments (De Jonghe and Rasmont 1983), enzyme electrophoretic data (Scholl and Obrecht 1983;Pamilo et al 1987) and analysis of the compounds of the secretion from male labial glands (Bertsch 1997) confirmed the existence of these species. Also, comparison of mtDNA sequences within the subgenus Bombus showed that species can be characterised using specific mtDNA polymorphisms (Pedersen 2002;Murray et al 2008). However, reliable identification based exclusively on morphological traits is still problematic and numerous discrepancies in classification and identification are still present.…”